Making sequencing simpler with nanopores

The Advances in Genome Biology and Technology (AGBT) conference, one of the main go-to destinations for those who get excited by DNA sequencing technology, is currently going down in Florida. Sadly, no-one from GNZ could make it this year, but we are keeping up with the various announcements about new genomics tech as best we can. One that caught our attention was the announcement of a brand new sequencing machine from a company that has previously kept very quiet about its technology.

A lot of the interest has come from the (very cool) MinION, a tiny, disposable USB-key sequencer (shown in the picture above) that can sequence about a billion base pairs of DNA, and cost around $500-$900 each. The applications of this are endless – the ability to pick up a bit of biological matter, mix it with a few chemicals, and read whatever DNA is in it, could help with diagnostics, epidemiology, ecology, forensics. It is also (though not quite) the price where hobbyists could consider having a play; perhaps in a few years plug-and-play DIY genetics could be a possibility.

Less immediately striking, but still just as interesting, is the GridION sequencing machine. This is the work-horse of the nanopore sequencing world, made for reading lots of DNA, and scaling up to massive sequencing centers. Obviously, many scientists are going to be very interested in many of the features (notably, the ability to read very long pieces of DNA, a trick that has previously been more-or-less impossible to do reliably). However, what will this announcement mean for those of us who are interested in personal genomics?

Making sequencing simpler

As a ball-park figure, one GridION machine (or “node”) will be able to read sequence at roughly 600 million base pairs per hour, which equates to around 14Gbp a day or a high coverage (30X) human genome in just under 6 days. A stack of 4 could thus produce sequence at about the same rate as one of Illumina’s top-of-the-range HiSeq 2500 machines. The cost will be similar as well, at around $2200-$3600 per genome. Data quality should be competitive (the company say that error rates are still higher than they’d like, but they are well on the way to getting to an accuracy of >99% by release), and the fact that it reads much larger pieces of DNA will get rid of many of the alignment errors that plague current generation sequencers.

In essence, the new machine should be able to compete with the best-in-class in terms of throughput, cost and data quality. This in-and-of itself is no small achievement for a small company, given the dominance of the large sequencing giants. If nothing else, introducing a radically different technology to the field will drive down costs. I wouldn’t be surprised if Oxford Nanopore was one of the first companies reliably producing $1000 genomes in 2013, but the the rate at which this field moves at makes predicting who will be sequencing DNA the cheapest even in six moths impossible.

However, what is really interesting about this new offering is that it manages to get around many of the niggling annoyances involved in sequencing DNA. The sequencers are very small, and stack neatly together – a stack of 20 is about the same size as a HiSeq, giving them a fifth of the footprint per genome sequenced. They use up a lot less power as well, needing about a sixth of the electricity per genome. The low cost of the machines themselves, and the plug-and-play nature of the system, means that scaling up is pretty simple too, and the fact that the machines can handle much of their own computing will keep down IT costs. Most importantly, getting the DNA ready for sequencing is a lot easier, and doesn’t need days of processing and expensive equipment required to prepare DNA for most machines. At AGBT, Oxford Nanopore CTO Clive Brown told a rather impressive story of sequencing DNA from rabbit blood, which involved bleeding the rabbit, diluting its blood in water, and sticking it into a tiny DNA sequencer attached to a laptop.

Basically, sequencing genomes will become a much simpler prospect in the near future, and in terms of providing sequencing to patients or to consumers, this can only be a good thing. As consumables costs for sequencing keep falling, the real cost of providing people with their genome sequencing is going to be dominated by the cost of floor space, of employing technicians, running computers, handling and preparing samples. I think that when we look back at what Oxford Nanopore did differently, no-one will remember that they sequenced a genome for 20% cheaper. They will remember that they used a laptop computer to sequence DNA straight from a rabbit.

What is it, exactly, that costs $2200-$3600? It has been my assumption that reagent cost was going to be negligible with nanopore sequencing, but if the cost is reagent cost then my assumption incorrect.

The cost is all-included: the array (which burns out and has to be replaced every few days), plus the reagents, and plus the machine (amortised over its life). I’d guess the biggest cost in all that is in manufactoring the array, so the main thing that will bring the price down is reducing the burn-out time for the electronics, or increasing the speed of the enzyme.

While the MinIon has gotten most of the attention, I haven’t yet figured out for what you might really use it. The problem is with the throughput, I have seen figures of about 160 Mb per hour, for six hours. So for a human genome you’d have to use up to ten of these devices, especially at 4% Indel Rate, and then the six hours are probably stochastic.

For genotyping we don’t really need 100Kb read lengths. But even if you chop up the dna, you’d still have to read a few gigabases to get some usefulamount of information for genotyping, again using up more than one device. Maybe running the sequencing at higher speed but lower quality might work for genotyping.

The one use I can think of in sequencing is bacterial sequencing. Much lower sizes of genomes, and most microbiologists until now don’t sequence that often.

Next use would be some kind of non-genetic diagnostics, maybe RNA, maybe Proteins. However, economics of scale would favor the GridIon system.

My understanding is that the nanopore devices use an array of nanopores or nanopore “units” at least, where each unit is able to measure its current independently. The limited running time probably comes from these pores degrading or breaking down. This should be a stochastic process, but over time the pores, and thus the sequencing speed decreases.

That will be true both for the MinION and for the GridION cartridges. Sequencing costs will thus depend a lot on the available time, patience, luck etc of the researcher.

The usecases of MinION will be pretty limited, at least at first. It might also be geared more towards protein diagnostics or even analysing pcr products.

For example you could sequence the pooled pcr products of a few hundred or thousands of individuals (flies, cancer cells etc), and just sequence them. Even microsatellite panels might be cost-effective.